Catherine E. Lofton-Day

Interleukin 21 and its receptor are involved in NK cell expansion and regulation of lymphocyte function.

Cytokines are important in the regulation of haematopoiesis and immune responses, and can influence lymphocyte development. Here we have identified a class I cytokine receptor that is selectively expressed in lymphoid tissues and is capable of signal transduction. The full-length receptor was expressed in BaF3 cells, which created a functional assay for ligand detection and cloning. Conditioned media from activated human CD3+ T cells supported proliferation of the assay cell line. We constructed a complementary DNA expression library from activated human CD3+ T cells, and identified a cytokine with a four-helix-bundle structure using functional cloning. This cytokine is most closely related to IL2 and IL15, and has been designated IL21 with the receptor designated IL21R. In vitro assays suggest that IL21 has a role in the proliferation and maturation of natural killer (NK) cell populations from bone marrow, in the proliferation of mature B-cell populations co-stimulated with anti-CD40, and in the proliferation of T cells co-stimulated with anti-CD3.

Identification of INSL6, a New Member of the Insulin Family That Is Expressed in the Testis of the Human and Rat

Abstract A new member of the insulin gene family (INSL6) was identified from an Expressed Sequence Tag database through a search for proteins containing the insulin family B-chain cysteine motif. Human and rat INSL6 encoded polypeptides of 213 and 188 amino acids, respectively. These orthologous sequences contained the B-chain, C-peptide, and A-chain motif found in other members of the insulin family. Human INSL6 was 43% identical to human relaxin H2 in the B- and A-chain regions. As with other family members, human and rat INSL6 had predicted dibasic sequences at the junction of the C-peptide and A-chain. Human INSL6 sequence had an additional dibasic site near the C-terminus of the A-chain. The presence of a single basic residue at the predicted junction of the B-chain and C-peptide suggests that multiple prohormone convertases are required to produce the fully mature hormone. INSL6 was found to be expressed at high levels in the testis as determined by Northern blot analysis and specifically within the seminiferous tubules in spermatocytes and round spermatids as detected by in situ hybridization analysis. Radiation hybrid mapping placed the human INSL6 locus at chromosome 9p24 near the placenta insulin-like homologue INSL4 and the autosomal testis-determining factor (TDFA) locus.

Molecular cloning and sequence analysis of the cDNA encoding human apolipoprotein H (beta 2-glycoprotein I).

SummaryApolipoprotein H, also known as β-2-glycoprotein I, was purified from human serum, and antiserum produced to denatured apolipoprotein H detected a cDNA clone from a γ gt11 library derived from human liver. This cDNA coded for the complete sequence of the mature protein. The cDNA insert, along with a polymerase chain reaction product which extended the 5′ end of the message, were subcloned and both strands were sequenced. The apolipoprotein H precursor was found to code for 345 amino acids, 326 of which appear in the mature protein. The deduced amino acid sequence of human apolipoprotein H differs from its rat homologue by the presence of a 48-amino acid stretch which is absent from the rat protein. The remainder of the proteins share a greater than 80% similarity. The amino acid sequence of apolipoprotein H consists largely of repeated units approximately 60 amino acids in length. These repeats are comparable to “sushi structures” found in a large number of diverse proteins, including complement components, receptors and regulators of complement activation, serum proteins, membrane-associated adhesion proteins, and other structural and catalytic proteins. Apolipoprotein H was shown to be transcribed by human hepatoma cell lines Hep 3B and Hep G2, and rat liver by detection of mRNA using northern blot analysis.

The genomics of Insulin 5

Advances in the development of sequence databases and novel bioinformatics algorithms have changed the way many new hormones are discovered. Nucleic acid sequence-based queries targeted to conserved features of the insulin/relaxin superfamily, have led to the discovery of a novel member of the relaxin family of hormones, INSL5 [1]. The discovery of a novel gene from DNA sequence permits the rapid analysis of gene expression patterns and chromosomal localization in an attempt to elucidate the normal physiological or pathological role of the hormone. However, the protein to gene discovery path practiced with all previously identified members of this hormone family benefited from the knowledge of the biologically active form of the hormone from the very onset, as well as a bioassay with which to follow the hormone. This is not the case with genomic discoveries since the signalling form of members of this hormone family are usually the product of multiple steps of posttranslational modification and for many members, the structure of the mature form is not obvious. In addition, the varied biological activities and unique organ and cellular location of the current members of the insulin/relaxin family make the elucidation of the biology of its orphan members a difficult task. The first steps toward our understanding of INSL5 biology has focused on experiments necessary to provide information on the structure of the mature hormone, the identification of some of the organ systems affected by its action, and the identification of a population of cells which make this hormone.